Previous studies on EBV-transformed B-cells indicated a classic pattern of susceptibility to HIV-1 that was governed by interaction of viral envelope with CD4 and the co-receptor associated with the tropism of the infecting strain. One exception involved dual-tropic strains, which are defined by the use of both CCR5 and CXCR4, and occasionally to a lesser degree, other co-receptors. We found that EBV B-cells derived from individuals homozygous for the CCR5-delta-32 (Null) genotype appeared to be less susceptible to infection with dual-tropic strains of HIV than would have been predicted by the absence of a functional CCR5 receptor on these cells. The studies during the present year have focused on expanding the panel of EBV-transformed B-cells from both CCR5 null and wild-type individuals; we show that there is a pattern of susceptibility to mono- and dual-tropic strains of HIV-1 that is dictated by the level of CD4 and co-receptor expression. Receptor expression on these B-cells approximates the minimum threshold level required for certain strains of HIV-1 to succeed in establishing productive infections; this appears to be especially true for dual-tropic strains. To confirm this observation, we have devised a molecular-based strategy for expanding our panel of dual-tropic strains of HIV-1 by testing a large bank of patient isolates. A second phase of this present study has been to investigate the susceptibility of primary non-transformed B lymphocytes to HIV-1 infection. Stimulation of CD40 with either soluble or membrane- bound ligand (CD154) was used to expand peripheral blood-derived B lymphocytes into cultures of high purity. Our findings indicate that as the cells proliferate under these conditions, there is increased expression of activation-related markers such as CD80, CD86 and CD54, as well as the HIV receptors CD4 and CXCR4; in contrast, CCR5 levels remained undetectable. Concomitantly, susceptibility to both T-tropic and dual-tropic strains of HIV also increased during the proliferation period. However, similar to the situation described for EBV-transformed B cells, we found that support for dual-tropic strains lagged behind T-tropic strains, suggesting that higher thresholds of CXCR4/CD4 double positive cells are required to support infection with dual-tropic strains. We have also directly established that HIV infects B lymphocytes through CD4 and we are currently addressing the dependence of this phenomenon on specific chemokine receptors.